The innate immune response represents the first barrier against viral infections and provides the initial signal required for development of an appropriate immune response. Interferon (IFN) constitutes a critical element of the innate immune system and is a current therapeutic agent that is effective against a broad spectrum of viral infections. However, new antiviral drugs are particularly needed for highly pathogenic viruses for which no vaccine is available or for virus strains resistant to viral inhibitors. Because the IFN response induces a cellular antiviral state that prevents virus replication, compounds that induce and/or activate the Interferon response will represent exciting candidates as therapeutics to treat viral infections. This application aims to identify interferon-inducing small molecule compounds for the treatment of a broad spectrum of viral infections and other interferon-related human diseases. These interferon-inducing compounds could be also used as immunomodulators and/or adjuvants for viral vaccines, illustrating their immediate and significant impact on human health. We will use a novel, simple, and sensitive cell-based assay to identify compounds that induce and/or activate the innate immune response. To achieve this, we will: develop and adapt a cell-based assay to identify compounds that induce and/or activate the innate immune response (Aim 1). Specifically, we will screen two libraries of structurally diverse small molecule compounds to demonstrate the feasibility of our cell-based assay to be translated to automated high-throughput screening (HTS) format. Next, we will use secondary assays to validate identified hits and their biological relevance (Aim 2). We will certify the compounds active in the primary screen as positive hits by secondary assays, evaluating their species specificity, mechanism of action and toxicity in order to prioritize their characterization in antiviral assays. Our straightforward cell-based assay represents a comprehensive and reliable approach for the immediate identification of promising IFN modulators to treat viral infections and other human diseases that rely on the unique use of IFN. This project will allow us to reach our long term goals of demonstrating the antiviral properties of the identified compounds in preclinical animal models to select lead therapeutic candidates. PUBLIC HEALTH RELEVANCE: There is a great need for new antiviral drugs, particularly against highly pathogenic viruses for which no vaccine is yet available or virus strains that become resistant to viral inhibitors. Because the interferon (IFN) response induces an antiviral state in cells that prevents virus replication, compounds that enhance or activate the IFN response represent lead candidates as antivirals, vaccine adjuvants, or potential immunomodulators. Using our high throughput cell-based assay, we will identify IFN-modulating compounds that will represent promising therapeutics to treat viral infections and other human diseases that rely on the unique use of IFN.